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Ion transport across posterior gills of hyperosmoregulating shore crabs (Carcinus maenas): amiloride blocks the cuticular Na⁺ conductance and induces current-noise

Horst Onken^1,* and Sven Riestenpatt²

¹ Departmento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Avenida Bandeirantes 3900, 14040-901 Ribeirão Preto, SP, Brasil and
² Biologische Anstalt Helgoland, Zentrale Hamburg, Notkestrasse 31, 22607 Hamburg, Germany

View larger version (16K): [in a new window]   Fig. 1. The effects of external amiloride (100 µmol l–1) on the fluxes of Cl– (A) and Na+ (B) across split lamellae of the posterior gills of shore crabs adapted to 10  salinity. Open columns, control values; hatched columns, after amiloride. Values are means + S.E.M. (N=5).

View larger version (13K): [in a new window]   Fig. 2. Representative time course of the uncorrected short-circuit current (I*sc) across split gill lamellae of shore crabs adapted to 10  salinity showing that ouabain (5 mmol l–1) completely inhibits the negative I*sc even in the presence of external amiloride (100 µmol l–1). The vertical current deflections are due to voltage pulses (1 mV) and directly reflect the (uncorrected) conductance of the preparation in the chamber.

View larger version (37K): [in a new window]   Fig. 3. Power density spectra obtained with split gill lamellae (A,B) or isolated gill cuticles (C,D) at different external amiloride concentrations (A,C; at a clamp voltage of 50 mV) and clamp voltages (B,D, at 5 µmol l–1 external amiloride) as indicated in the insets. Sf, power density; f, frequency.

View larger version (15K): [in a new window]   Fig. 4. Hanes–Woolf plots (concentration of amiloride, cAmi, versus ratio of cAmi to the induced change in Isc as a percentage of the control current, EAmi) of the influence of external amiloride on the currents across split gill lamellae at a clamp voltage of 50 mV (circles; mean 100 % control current=3635±475 µA cm–2) and on the currents across isolated cuticles at clamp voltages of 50 mV (squares; mean 100 % Icut=28 530±4889 µA cm–2) or 10 mV (triangles; mean 100 % Icut=4782±910 µA cm–2). Icut, cuticular current. The lines correspond to linear regressions (r2>0.99 in all cases) over the entire amiloride concentration range used (1.25–200 µmol l–1 for split gill lamellae and 0.5–100 µmol l–1 for isolated cuticles). Values are means ± S.E.M. (N=5 in all cases).

View larger version (15K): [in a new window]   Fig. 5. The dependence of the mean Lorentzian plateau values (So) on increasing concentrations of external amiloride for split gill lamellae (circles; N=5) and isolated cuticles (squares; N=3) at a clamp voltage of 50 mV. Values are means ± S.E.M.

View larger version (14K): [in a new window]   Fig. 6. The dependence of the mean values of 2fc on increasing concentrations of external amiloride for split gill lamellae (circles; N=5) and isolated cuticles (squares; N=3) at a clamp voltage of 50 mV. Values are means ± S.E.M. fc, corner frequency.

View larger version (14K): [in a new window]   Fig. 7. The results of calculations of the ‘channel’ density (squares; N=3; mean + S.E.M.) and the single-‘channel’ current (circles; N=3; mean - S.E.M.) for isolated gill cuticles at a clamp voltage of 50 mV. Values are means ± S.E.M. i, single-‘channel’ current; M, the number of ‘channels’ per cm2.